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Understanding Antibiotics Mechanism in SynBio: Ampicillin, Kanamycin, and Chloramphenicol

Explore how antibiotics like ampicillin, kanamycin, and chloramphenicol work in synthetic biology, the resistance mechanisms, and their applications. Learn about their bactericidal and bacteriostatic effects, target specificities, and implications for genetic purposes.

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Understanding Antibiotics Mechanism in SynBio: Ampicillin, Kanamycin, and Chloramphenicol

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  1. The mechanism of antibiotics Biol 1220 Synthetic Biology abe pressman & minooramanathan

  2. the basics • Used to kill or inhibit the growth of bacteria • Classified as bactericidal or bacteriostatic Kill bacteria directly Prevent cell division • Classified by target specificity: Narrow-spectrum vs Broad range • Most modified chemically from original compounds found in nature, some isolated and produced from living organisms

  3. sites of antiobiotic action

  4. ampicillin • Belongs to β-lactamgroup of antibiotics – contain β-lactam ring • Broad-spectrum • Penicillin derivative that inhibits bacterial cell wall synthesis (peptidoglycan cross-linking) • Inactivates transpeptidases on the inner surface of the bacterial cell membrane • Bactericidal only to growing E. Coli • Widespread use leads to bacterial resistance. HOW?

  5. ampicillin resistance • Cleavage of β-lactam ring by β-lactamase enzyme

  6. ampicillin resistance • β-lactamase is encoded by the plasmid-linked bla (TEM-1)gene • Hydrolyzes ampicillin • Ampicillin levels in culture continually depleted

  7. use in synthetic biology • To confirm uptake of gene (eg. of plasmids) by bacteria • Bacterial Transformation: DNA integrates into bacteria’s chromosome and made chemically competent • Exogenous DNA tagged with an antibiotic resistance gene eg. β-lactamase • Grown in medium containing ampicillin • Ampicillin resistance indicates successful bacterial transformation

  8. Kanamycin • Targets 30s ribosomal subunit, causing a frameshift in every translation • Bacteriostatic: bacterium is unable to produce any proteins correctly, leading to a halt in growth and eventually cell death

  9. kanamycin use/resistance • Over-use of kanamycin has led to many wild bacteria possessing resistance plasmids • As a result of this (as well as a lot of side effects in humans), kanamycin is widely used for genetic purposes rather than medicinal purposes, especially in transgenic plants • Resistance is often to a family of related antibiotics, and can include antibiotic-degrading enzymes or proteins protecting the 30s subunit

  10. chloramphenicol • Bacteriostatic: functions by halting bacterial growth, which is done by inhibiting the enzyme peptidyl transferase, a protein that assists in the binding of tRNA to the 50s ribosomal subunit • Three methods of resistance: reduced membrane permeability, mutation of the 50s subunit, and an enzyme called chloramphenicol acetyltransferase, which inactivates chloramphenicol by covaltly linking groups • Easy/cheap to manufacture, but unused in western countries because of possible aplastic anemia as a side effect

  11. Sources • http://en.wikipedia.org/wiki/Antibiotic • http://en.wikipedia.org/wiki/Ampicillin • http://en.wikipedia.org/wiki/Beta-lactamase • http://www.sigmaaldrich.com/catalog/ProductDetail.do?N4=A1593|SIAL&N5=SEARCH_CONCAT_PNO|BRAND_KEY&F=SPEC • http://abe.leeward.hawaii.edu/Protocols/QiagenSpinprepProtocol.htm • http://www.openwetware.org/wiki/Brown_BIOL1220:Notebook/SynBio_in_Theory_and_Practice/Bacterial_Basics

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